The DNA of a cell presents the first line of control over the metabolism and regulatory processes of the cell. However, the mechanisms by which the genetic message of a cell is maintained and transmitted to offspring are unknown. The proposed research will utilize genetic and biochemical techniques to study the mechanisms of DNA replication. We will make use of both bacterial and mammalian mutants defective in replication in these studies. A major portion of the effort will take advantage of in vitro systems of DNA replication. We will attempt to delineate the role of the DNA polymerases in replication. For this both bacterial mutants deficient in DNA polymerases and highly purified DNA polymerase will be used to define the capabilities of the enzymes and the possibility of overlapping roles. The appearance of intermediates of replication in mutants deficient in a DNA polymerase will be one aspect of this portion of the work. The appearance of new or altered forms of DNA polymerases in the case of reversion will be an important point of interest. In addition the control processes of initiation and termination of DNA replication will be studied using in vitro systems. Of particular interest here will be the function of the dnaA gene product and its role in initiation. We will attempt to differentiate between positive and negative regulatory models of control for initiation of replication. We will also investigate specific enzymes in several mammalian cell lines with defects suggestive or deficiencies in steps of replication. If the chromosome of the cell contains degenerate, uninterpretable or deleterious information, then the functional capacity of the cell will be impaired. Such defects may be the basis of cancerous changes in mammalian cells, structural defects in heighr organisms, or metabolic aberrations in either prokaryotes or eukaryotes.